With regard to the SPR measuring method which is particularly—but not exclusively—of interest in view of implementing the proposals of the invention, reference may for example be made, concerning the technical background, the metrological equipment and relevant applications, to: A. Zybin et al., Anal. Chem., 2005, 77, 2393-2399; T. Akimoto et al., Biosensors and Bioelectronics 2003, 18, 1447-1453; A. K. Sharma et al., Optics Communications 2005, 245, 159-169; C. E. H. Berger et al., Sensors and Actuators B 2000, 63, 103-108; H. B. Lu et al., Sensors and Actuators B 2001, 74, 91-99; S. A. Zynio et al., Sensors 2002, 2, 62-70; J. R. Lakowicz, Analytical Biochemistry 2004, 324, 153-169 und N. Nath et al., Anal. Chem. 2002, 74, 504-509. Reference may also be made to a differential surface plasmon resonance measuring device and a corresponding measuring process according to EP 1617 203 A1 (T. Imato et al.).
A good overview of the basic principles of SPR sensors, types of implementation of SPR biosensors and applications of SPR biosensors is provided by various authors in the textbook “Surface Plasmon Resonance Based Sensors”, Springer Series on Chemical Sensors and Biosensors (Ed. O. S. Wolfbeis), Volume 04 (Ed. J. Homola), Springer Verlag Berlin Heidelberg 2006.
An overview of developments and development objectives pursued in SPR biosensors is provided by Charles T. Campbell in an article entitled “Surface Plasmon Resonance (SPR) Biosensor Development” which can be downloaded from Internet on the following URL: www.cpac.washinqton.edu. This article also addresses differential SPR measurements in array format by means of what is known as SPR microscopy in which a 10×12 array is used with sixty measurement spots which are functionalized for binding sixty different biopolymers and sixty reference spots for subtracting background changes, in particular owing to non-specific binding and changes in the index of refraction of the buffer solutions.
The invention relates in particular to a technical solution in the field of analytical technology for physical, chemical and biochemical analysis. A particularly relevant application is, for example, determining the concentration of various substances in gaseous and liquid environments (cf. for example Class G01 N 21 of the International Patent Classification (IPC)), the carrying-out of biochemical analyses and immunological tests in medicine and in research, in biotechnology, for checking food quality, agricultural products and drinking water; this also includes determining the content of harmful substances (pesticides, insecticides, etc.), and also for the ecological monitoring of the environment. It may also involve the examination of adsorption processes, the highly sensitive measurement of temperature-dependent optical and electrical properties or examination of the change in these properties.
For the measurement of physical, biological, biochemical or chemical parameters, use is currently made above all of either direct or differential measurements. In the case of direct measurement, a measuring signal is read directly from an individual sensor. In the case of differential measurement, the signal is measured by an “active” sensor (also known as a measurement spot) and the signal, which is measured on an inactive sensor (also known as a reference spot), is subtracted from this measuring signal. Differential measurement allows the possible influence of a large number of disturbing factors and boundary conditions which it is impossible to check precisely, such as for example pressure, concentration of reagents, temperature and other parameters, to be prevented. The differential measuring assembly is well known and is used widely in technology and science. To give just a few examples: In physical sensors, differential measurements allow the influence of temperature to be minimized; in chemosensors and biosensors, the use of differential measurement allows not only the influence of temperature to be reduced but also non-specific effects on the surface of the sensor (for example adsorption of an interfering substance in affinity sensors, fluctuation of the oxygen partial pressure in enzymatic biosensors comprising a Clark electrode) to be compensated for. The use of differential measurements has been described in a large number of sources, in scientific publications and patent publications.
Nowadays, the sensitivity of many types of physical and chemical sensors is limited by fluctuations in temperature or fluctuations in the concentration of reagents or other physical and chemical parameters between the measurement spot and reference spot [A. Zybin et al., Anal. Chem., 2005, 77, 2393-2399]. To reduce these effects, certain measuring apparatuses use highly precise temperature stabilization. Chemical sensors and biosensors use microfluidics which are relatively complex and not always reliable to reduce the fluctuations in temperature after addition of reagents, thus minimizing the difference in temperature between the reagents added and the surface of the sensor. In this case too, the fluctuations in temperature are not compensated for completely, as the interaction of the analyte with the measurement spot can take place as an exothermic or endothermic reaction. Additional difficulties which are not compensated for by highly precise temperature stabilization and microfluidics include fluctuations in the concentration of reagents in addition to the surface area or other physical parameters (for example pressure, reagent flow rate, etc.).
To improve compensation for disturbing factors, the measurement spot and the reference spot should be placed as close as possible alongside one another. However, this is not possible without limitation: although the measurement spot and the reference spot can be placed on a surface almost without a spacing, the measuring signals supplied by a measurement spot or reference spot are for most measuring methods integral signals of whole measurement surfaces. If there is a temperature gradient or a reagent gradient on the surface of the sensor (and inevitably there will be), signals from measurement spots and reference spots will be roughly the same as the signals at the centers of the spots. Therefore, not only the spacing between reference spots and measurement spots but also the size of the spots will characterize the minimum difference in temperature thereof. The size of the measurement spots and reference spots can be reduced only to a limited extent, as this typically leads to a reduction in the signal/noise ratio.
With regard to the nomenclature, it should be noted that the terms “measurement spot” and “sensor spot” are frequently—by way of distinction from the associated reference spot—used synonymously, although strictly speaking the reference spot can also be identified as a sensor spot, namely as a sensor spot of a type other than the measurement spot.
The invention is based on the object of preventing or at least greatly reducing the problems of the prior art without necessarily incurring high equipment costs.